The diagnosis, prognosis, and therapeutic evaluation of acute neurodegenerative conditions such as traumatic brain injury, stroke, and cardiac arrest-induced brain damage would be aided considerably by minimally invasive surrogate measures for brain damage, but currently these are lacking. My laboratory has devised a neurobiological approach for identifying new surrogate markers for acute neurodegeneration measurable in accessible body fluids. We initiated the first large-scale analysis of protein release from degenerating cultured rat cortical neurons, and used an existing antibody collection along with methods of 2D PAGE and mass spectrometry to detect over 60 polypeptides released preferentially from degenerating neurons. The potential of the approach for identifying surrogate markers for brain injury in vivo has been established by antibody-based detection of released polypeptides in cerebrospinal fluid or serum following experimental traumatic brain injury or cerebral ischemia in rats, as well as thoraco-abdominal surgery with circulation arrest in humans. The proposed research would extend the preclinical and clinical applications of these findings. Specific Aim 1 will identify neuronal cytoskeletal proteins and their proteolytic fragments that are released preferentially by degenerating cultured neurons. Specific Aim 2 will define the proteomes for protein release triggered by necrotic and apoptotic neurodegeneration. Novel antibodies and sandwich immunoassays will be developed for the sensitive and specific quantitation of potential markers emerging from these two aims. Specific Aim 3 will evaluate the markers by immunoassays of cerebrospinal fluid, serum, and urine samples derived from an experimental model for traumatic brain injury, and establish relationships between time-and treatment-dependent changes in marker levels and brain histopathologies. Specific Aim 4 will develop immunoassays for human biomarkers and examine them in a study of human head injury. The overall goal of the project is to identify novel surrogate markers for acute brain damage measurable in accessible body fluids, develop immunoassays enabling their quantitation, and begin to assess their preclinical and clinical utilities as diagnostics, prognostics, and surrogate measures of neuroprotectant efficacy.